Natural L-type calcium channels antagonists from Chinese medicine

L-type calcium channels (LTCCs), the largest subfamily of voltage-gated calcium channels (VGCCs), are the main channels for Ca2+ influx during extracellular excitation. LTCCs are widely present in excitable cells, especially cardiac and cardiovascular smooth muscle cells, and participate in various Ca2+-dependent processes. LTCCs have been considered as worthy drug target for cardiovascular, neurological and psychological diseases for decades. Natural products from Traditional Chinese medicine (TCM) have shown the potential as new drugs for the treatment of LTCCs related diseases. In this review, the basic structure, function of LTCCs, and the related human diseases caused by structural or functional abnormalities of LTCCs, and the natural LTCCs antagonist and their potential usages were summarized.

LTCCs, the largest subfamily of VGCCs, are the main channel of Ca 2+ influx in the cell excitatory process, which closely related to excitation-contraction coupling (ECC) and excitation-secretion coupling [3].LTCCs are widely present in various excitable cells, especially cardiac and cardiovascular smooth muscle cells, which are essential for heart and nervous function [3].Cav1.1 of LTCCs is distributed in skeletal muscle, and its mutation is associated with hypokalemic periodic paralysis type 1 (HPP-1) [4] and malignant hyperthermia [5].Cav1.2 and Cav1.3 are primarily existed in the heart and brain, and are related to Timothy syndrome (TS) [6,7], arrhythmia, bipolar disorder (BD) [8] and autism [7].Cav1.4 is presented in the retina and variants of Cav1.4 lead to night blindness [9].
General speaking, regulation of LTCCs has been considered as an important strategy for treating diseases for decades.LTCCs antagonists have been used for the treatment of hypertension, arrhythmia and other diseases, which illustrated their therapeutic activities in myocardial ischemia protection (MI), myocardial and vascular wall hypertrophy prevention, atherosclerosis prevention, and renal protection [10,11].Clinical-used LTCCs antagonists can be divided as 1, 4-dihydropyridine, benzothiazole, and phenylalkyl amine according to their chemical structures [12].
Traditional Chinese medicine (TCM) has a long history in cardiovascular diseases, among which Salvia miltiorrhiza, Ligusticum wallichii, Angelica sinensis, Paeonia lactiflora and Paeonia suffruticosa exhibited the calcium antagonistic effect [13].A total of 45 active ingredients from Chinese medicine with antihypertensive effect were screened though pharmacophore model based on drug repositioning method, which suggested that the Chinese medicine were the potential source of LTCCs antagonists [14].Therefore, it is of great value to develop and design efficient LTCCs antagonists from TCM.
In this review, we summarized the basic structure and molecular functions of LTCCs, related diseases caused by channel dysfunction.In addition, the LTCCs antagonists with different types from natural products of TCM were also simiply summarized.Furthermore, we hope to discover new natural LTCCs antagonists with high specificity in treating human diseases.
Cav1.4 is largely localized in the retinal cells and affects the release of neurotransmitter and Cav1.4 influences the dihydropyridine-sensitivity without the Ca 2+ currents [9].
The auxiliary subunits β belongs to the MAGUK-scaffolding protein family, a cytosolic soluble protein with high affinity binding to channel, including four subtypes of β 1 -β 4 [16].The mutation of β subunit is associated with arrhythmia and stroke [16].The α2δ subunits are encoded by one unique gene, and translationed into two separate proteins that linked by disulfide bond.The four subtypes of α2δ proteins, α2δ 1 -α2δ 4 , function as a thrombospondin receptor to regulate excitatory synpatogenesis [2,16].There are eight subtypes of γ subunit (γ 1 -γ 8 ), that is composed of four transmembrane helical segments with intracellular amino (NH2) and carboxy (COOH) termini.The physiological function of the γ subunit needs further research to reveal [2,16].The complex structure of Cav1.1 (α1, α2δ, β, and γ) from rabbit skeletal muscle membranes were determined by cryo-EM, which layed foundation for further understanding the working mechanisms of LTCCs with important physiological and pathological functions [17,18].

Cardiovascular disease
Ca 2+ is involved in many cellular processes such as EC coupling, membrane excitability and transcriptional regulation of cardiomyocytes through LTCCs influx, and plays an important role in physiological functions of cardiomyocytes [11].The Cav1.2, Cav1.3 and auxiliary subunits, including β subunits, α 2 δ subunits in myocardium, participate in the regulation of the activation and inactivation characteristics of the channels [11,19].The disorder of LTCCs can lead to electrophysiological abnormalities, arrhythmias, and various Ca 2+ dependent dysfunctions in cellular processes.
Myocardial ischemia/reperfusion injury (MI/RI) refers to the severe injury of ischemic myocardial tissue after restoration of perfusion and the clinical manifestations include reperfusion arrhythmia, intracardial hemorrhage and myocardial infarction [20].Calcium homeostasis is essential for maintaining ECC in cardiomyocytes, including calcium release, recapture, and storage [10].Studies have shown that MI/R injury can cause disruption of calcium homeostasis and Ca 2+ overload is one influencing factors in MI/RI, which may lead to energy metabolism disorder, myocardial hypoxia and ultimately myocardial cell necrosis [21].The regulatory proteins, including LTCC, provide potential targets for the prevention and treatment of clinical MI/RI.Hypertension is closely associated with increased vascular contraction.The influx of Ca 2+ into the vascular smooth muscle cells produce membrane potential and increase the tension of the blood vessel, which affect the arterial contraction and blood pressure [22].Moreover, the calcium sensitization process in smooth muscle cells could increase the vascular smooth muscle contraction though DAG-PLC-PKC pathway and the RhoA-Rho kinase pathway [23].
The mutation of the LTCCs causes the imbalance of Ca 2+ in cells and the changes of membrane potential, which causing the myocardial cells to be unusually excited and eventually lead to cardiac dysfunction [19].The mutations of α 1 C and β 2b may lead to Idiopathic ventricular fibrillation.The absence of CACNA1C p.E850 may reduce the surface expression of LTCC, which leading to a significant reduction of I Ca .Inactivation of Cav1.3 bring out a strong decrease of I Ca in the sinoatrial nodal pacemaker cells, which resulting in sinoatrial node dysfunction manifested as sinoatrial arrhythmia and bradycardia [24].Brugada syndrome (BrS) is an inherited arrhythmia related to mutations of 18 different genes, of which α 1 C mutation ranked the second common cause.In particular, CACNA1C and CACNB2 mutations accounted for 12% of BrS cases.The latest study identified two BrS-related mutation sites between domains I and II of Cav1.2, including p.T320M and p.Q428E [25].

Neurological disorders
LTCCs are also essential for neuronal functions.The mutations in LTCCs genes have a close relationship with various neurological and psychiatric disorders, including Timothy syndrome (TS), Parkinson's disease (PD), Alzheimer's disease (AD), epilepsy, Tuberous sclerosis complex (TSC) and drug addiction.
Timothy syndrome (TS) is a debilitating and multiorgan disease involving mental retardation, fatal arrhythmias and autism [6,7].Studies revealed that Cav1.2 channel mutations in TS patients leaded to impaired neural circuits [26].In addition, genome-wide association studies have found significant associations between α 1 C intron SNPs and psychiatric disorders including BD, schizophrenia and autism spectrum disorders [6].The characteristics of typical and atypical TS phenotypes have been summarized in a recent review to elucidate the molecular mechanism of Cav1.2 gated dysfunction leading to mental illness [27].
Dysregulation of calcium homeostasis is a compensatory result of neurodegenerative processes in early Parkinson's disease (PD).The amount of Cav1 subtypes and the calcium-binding proteins in PD were different from control group.The increased expression of Cav1.3 subtype in the cerebral cortex of early stage PD patients may subjoin the cells metabolic burden that depend on LTCCs subtypes for electrical activity, which lead to the neurodegeneration of specific groups of neurons.The change in ratio of Cav1.2 to Cav1.3 in parkinsonian brain could increase neuronssusceptible to excitotoxicity or oxidative stress [28].
Alzheimer's disease (AD) is characterized by the accumulation of β-amyloid peptide (Aβ), dysfunction of synapses, and loss of neurons.The increasement of age associated oxidative stress and metabolic disorders cause dysplasia of calcium homeostasis.The extracellular accumulation of Aβ enhance calcium load and increase the current of the Cav1.2 and Cav1.3 in AD [29].Moreover, blocking calcium channels alleviate amyloid-β-induced neuronal decline in vitro and exhibited neuroprotective effects [30].
Epilepsy refers to the recurrent brain dysfunction resulted from sudden excessive and disordered neuronal discharge.Nimodipine can block abnormal spontaneous activity of hippocampal pyramidal neurons of heat-induced in Cav1.2 knock-out mice brain slices, suggesting that Cav1.2 subunit is critical in temperature-dependent intrinsic firing of febrile epilepsy [31].
Tuberous sclerosis complex (TSC) is neurologic impairment that associated with epilepsy.The development of TSC epilepsy closely related with high activity of TSC2-deficient (TSC2 −/− ) neurons.The specific inhibitor of mTOR protein, rapamycin, could inhibit the abnormal increase of LTCCs subtype Cav1.3 in TSC2 −/− neurons.The results indicated that LTCCs were critical downstream component of TSC-mTOR signal and can trigger the enhancement of network activity of TSC2 −/− neurons [32].Therefore, LTCCs may be a new therapeutic target for TSC epilepsy.
Drug addiction, known as drug dependence or drug abuse, is a stubborn and chronic recurrent neurological disease.Ca 2+ ions and Ca 2+ channels are involved in the formation of drug addiction, and L-type Ca 2+ channels are an important target for anti addiction drug research [33,34].Studies showed that Cav1.2 channels, rather than Cav1.3, are involved in withdrawal reaction in nicotine-induced abuse and alcohol-seeking abuse [35,36].LTCC blockers, such as dihydropyridines, have been considered a potential therapeutic drug to alleviate the symptoms of drug addiction [37].

Psychological diseases
Several studies have implicated that LTCCs disorders may lead to psychiatric ills, such as BD, and schizophrenia [7,9,19], which suggested the importance of LTCCs in learning, memory, and synaptic plasticity.
The Genomic data suggested that CACNA1S, CAC-NA1C and CACNA1D were the core genes that related with psychiatric diseases.Calcium signaling dysfunction is one pathogenic factor for psychological diseases [38][39][40][41].The subunit α 1 C, α 1 B and β 2 subunits were risk locus for BD, schizophrenia and recurrent major depression [38,42].The microRNA 137 has been proved as a potentially risk for schizophrenia, and the CACNA1C, one target of microRNA 137, influenced the development process of schizophrenia [39].
LTCCs antagonists has been used for the treatment and prophylaxis of psychological diseases over 30 years, but has not turned into an established therapeutic scheme [8].The results of existing studies about LTCCs antagonists on other phases of the illness were limited to the observational research without robust evidence.However, long-term or excessive usage of LTCC antagonists increased the risk of cardiovascular disease and depression [40].Thus, it is important to comprehensive utilize the pharmacological, molecular, and genetic material to ameliorate the efficacy, safety and selectivity of LTCCs antagonists in the clinical treatment of psychological diseases.

Aldosterone-producing adenomas (APAs) APAs account
for approximately 50% of primary aldosteronism, which is one of the most common causes for hypertension patients.Zona glomerulosa (ZG)-like APAs have four somatic mutations, V259D, G403R, I750M and P1336R, in the Ca 2+ pore at the S5 and S6 domains of Cav1.3 [43].Selective Cav1.3 blockers may treat ZG-like APAs hyperaldosteronism without the vascular side effects caused by current LTCCs blockers.
Congenital hearing impairment Congenital hearing impairment has extensive genetic heterogeneity.Cav1.3 is expressed in cochlear hair cells and is critical for auditory brainstem development [9].α 1 D mutations in Cav1.3 was found in two consanguineous families with deafness and severe mouse sinoatrial node dysfunction with bradycardia [44].
Congenital stationary night blindness type 2 (CSNB2) CSNB2 patients exhibit some degrees of night blindness, low visual acuity and myopia [45].Cav1.4 of LTCCs is mainly expressed in retinal neurons, especially at the photoreceptor terminals.The mutations in the CACNA1F gene that encodes Cav1.4 channels lead to the the channel activity altered and caused the retinal disease, for example, CSNB2 [9].The different structural, functional phenotypes and treatment options of Cav1.4 mutations in CSNB2 were summarized in recent review [46].The gene therapeutic maybe a promising approach to CSNB2 patients in future.

LTCCs antagonists from natural products
LTCCs are implicated in multiple cardiovascular, neurological and psychological diseases, and has become an important target for drug development.Natural products have been considered as valueable sources for drug discovery as their fewer adverse effects and multiple mechanisms.Many TCM active ingredients have been reported with inhibitory effect on LTCCs through various mechanisms (Table 2), including reducing the expression of Cav1.2 and Cav1.3 subunits and related proteins, inhibiting calcium channel currents, restricting calcium influx, and decreasing calmodulin-dependent protein kinase II (CaMKII) signaling pathways (Fig. 2).

Polyphenols Paeonol
Paeonol (1, Fig. 3) is an active polyphenol from the root bark of Paeonia suffruticosa Andr.In previous study, paeonol exhibited protective effect on acute myocardial infarction rats by inhibiting LTCCs currents in rat ventricular myocytes [47,48].In addition, paeonol induced non-endothelium dependent-vasodilatation in rat mesenteric artery by inhibiting VGCCs via inducing extracellular Ca 2+ influx [49].Therefore, the mechanism of paeonol in reducing myocardial infarction and protecting myocardial cells from hypoxia injury may be related to inhibition of LTCCs.In another study, paeonol alleviated primary dysmenorrhea by inhibiting Ca 2+ influx and uterine contraction via cannabinoid receptor 2 (CB2R) in uterine smooth muscle cells through MAPK/ERK pathway.As a result, paeonol exhibited the similar effect as positive control, nifedipine, in suppressing uterine contraction in vitro [50].

Salvianic acid A and salvianolic acid B
Salvianic acid A and salvianolic acid B (2-3, Fig. 3) are the main water-soluble constituents from Salviae Milthiorrhizae Bge., which have been used to treat cardiovascular diseases such as stroke, myocardial infarction and hypertension [51].Salvianolic acid A and salvianolic acid B exerted cardioprotective effects by reducing L-type Ca 2+ channel currents (I Ca,L ), upshifting the current-voltage, leftshifting the curves of activation and inactivation, and inhibiting the amplitude of the cell shortening [52,53].Moreover, salvianolic acid B showed vasorelaxant effects on isolated coronary artery rings precontracted with 5-hydroxytryptamine by inhibiting Ca 2+ influx in the vascular smooth muscle cells [54].

Flavonoids Luteolin
Luteolin (4, Fig. 4) is a natural flavonoid isolated from many traditional medicines and has various pharmacological activities in osteoporosis, allergy, diabetes, tumors and liver toxicity [55].The large amount of oxygen free radicals and calcium overload in myocardial cells are the main causes of heart injury.Furthermore, calcium influx through LTCCs during ischemia and hypoxia lead to further overload of calcium storage.Luteolin showed protective effect on heart from long-term preservation damage, such as structural damage, heart dysfunction and increased apoptosis by inhibiting hypoxia-dependent L-type calcium channels, which suggested the usage of luteolin as heart preservation solutions, especially in long-term storage [56].

Calycosin
Calycosin (5, Fig. 4) is an isoflavonoid isolated from Radix Astragali (Astragalus membranaceus (Fisch.)Bunge.), and showed the vasorelaxant effect on denuded and intact endothelium aortic rings [57].Calycosin reduced aortic ring contractions induced by agonists KCl     and PHE and its vasorelaxant action was different from that of dihydropyridines.The study revealed that calycosin was a noncompetitive Ca 2+ channel blocker that its vasorelaxant effect was endothelium-independent and was unrelated to intracellular Ca 2+ release [57].

Puerarin
Puerarin (6, Fig. 4) is an isoflavone C-glucoside isolated from Pueraria lobata (Willd.).The increased intraocular iron stimulates ROS generation lead to retinal injury.ROS showed the biphasic effect on the Ca 2+ transport in cells: on one side, ROS reduced the Ca 2+ -ATPase of sarcoplasmic reticulum (SR), which can diminish the Ca 2+ level in the cell; and on the other side, ROS deactivated the plasma membrane Ca 2+ -ATPase, which increase the Ca 2+ -loading and ultimately leads to apoptosis [58].Puerarin was reported the protection effect against retinal injury caused by iron overload though reducing the Cav1.2 expression in retinal tissue [59].Puerarin inhibited the level of Cav1.2 expression in ARPE-19 cells, indicating that puerarin attenuated the iron deposition by regulating the iron-handling proteins [59].The above results suggested that LTCC might be a potential target for puerarin on iron-mediated retinal injury.However, the specific target of puerarin inhibiting Cav1.2 needs further study.

Scutellarin
Scutellarin (7, Fig. 4) is a flavonoid glycoside from Erigeron breviscapus Hand Mazz.It had been reported that LTCC was activated by CaMKII, and CAMKII-mediated changes in calcium current may be associated with cardiovascular disease [60].The Ca 2+ -calcineurin and CaMKII were two important effector of Ca 2+ -medicated cardiac hypertrophy [61].In this study, scutellarin suppressed the cardiac hypertrophy exposed to phenylephrine (PHE) by inhibiting the Ca 2+ -mediated calcineurin and CaMKII pathways [61].Accordingly, scutellarin may be used as candidate against cardiac hypertrophy in future.

Hydroxysafflor yellow A
Hydroxysafflor yellow A (8, Fig. 4) is a water soluble constituent from Carthamus tinctorius L. and exerts various effects on cardiovascular diseases [62].Hydroxysafflor yellow A showed the cardioprotective effect on HR-induced myocardial injury in neonatal rat primary cardiomyocytes (NPCMs) and human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs).Further study showed that hydroxysafflor yellow A attenuated the expression of α1C and α2δ subunits of LTCC in vivo and in vitro.Bay-K8644, an LTCC agonist, was used to stimulate the LTCC excessive in study.As a result, Hydroxysafflor yellow A inhibited the electrical signal disturbances and the higher calcium currents caused by the excessive activation of LTCC in hiPSC-CMs, suggesting that hydroxysafflor yellow A treated MI/RI via regulating LTCC to inhibit Ca 2+ overload and apoptosis [63].

Paeoniflorin
Radix Paeoniae Alba, the root of Paeonia lactiflora Pall, has the effect of relieving depression and regulating menstruation in Chinese medicine.Paeoniflorin (10, Fig. 4) is the main bioactive terpene glycoside of paeony extract and has anti-depressive and neuroprotective effects.
It was confirmed that paeony extract and Shuyu capsule improved the depressive hehaviour, such as body weight, open-field test scores, and sucrose preference in premenstrual syndrome (PMS) rats by regulating Cav1.2 mediated CaM/CaMKII signalling.Further results revealed that paeoniflorin inhibited intracellular Ca 2+ overloading induced by K + and inhibited Cav1.2 current density in a dosage-dependent manner [65].Thus, paeoniflorin played an antidepressant role by mediating LTCCs.

Ginsenosides
Ginsenosides are the major bioactive ingredients from Panax ginseng.The total ginsenosides (TG) displayed cardioprotective effects on ISO-induced MI rats by inhibiting of I Ca-L , myocytes shortening and Ca 2+ transient [66].Ginsenoside Rb 1 and ginsenoside Rd (11-12, Fig. 5) have been reported the anti-MI effect in rat ventricular myocytes by inhibiting L-type Ca 2+ current in a dosagedependent manner [67][68][69].Ginsenoside Rb 1 and Re (13, Fig. 5) inhibited the mRNA expression of Cav1.2 on rat cardiomyocyte injury induced by aconitine alkaloids [70].Further study revealed that the inhibition of I Ca,L induced by ginsenoside Rd was abolished by pertussis toxin, a Gi protein inhibitor, suggesting that Gi protein was the potential target of Rd for treating MI/RI in rat [69].

Glycyrrhizic acid and its derivatives
Glycyrrhizic acid (14, Fig. 5) is a bioactive component isolated from Glycyrrhiza uralensis Fisch with wide range of pharmacological effects such as antiviral, antitumor, anti-inflammatory, bactericidal, and anti-MI [71].The neuroprotective mechanisms of glycyrrhizic acid was verified to be related with modulating multiple anti-apoptotic and pro-apoptotic factors and inhibiting intracellular Ca 2+ overload [72].Glycyrrhizic acid exhibited the cardioprotective effects by inhibiting L-type Ca 2+ channels and reducing the Ca 2+ transient in ISO-induced myocardial ischemia injury rats [73].Specifically, Glycyrrhizic acid decreased the elevation of ST segment, reduced the heart rate, increased the QT-interval shortening induced by ISO, and amended the heart morphology.Furthermore, Glycyrrhizic acid blocked L-type Ca 2+ channels in a dose-dependent manner and reduced the Ca 2+ transient in the rats ventricular myocardial cells [73].
The derivatives of glycyrrhizic acid also have similar biological functions.It has been reported that the calcium antagonists can suppress I Kr in ventricular myocardial cell that causes long QT syndrome (LQTS), which was a serious disease with a high risk of developing cardiac arrhythmias [74].An isomerized derivatives of glycyrrhizic acid, Magnesium isoglycyrrhizinate (15, Fig. 5), exerted cardiovascular protective effect by restraining I Ca-L and inhibiting Ca 2+ transient and decreasing myocardial contractility [75].In addition, Magnesium isoglycyrrhizinate showed no effect on the expression of I Kr in HEK293 cells, indicating that the usage of Magnesium isoglycyrrhizinate may not bring out drug-induced LQTS [75].Monoammonium glycyrrhizinate (16, Fig. 5), an ammonium salt hydrate of glycyrrhizic acid, often clinically applied in treating viral hepatitis.Monoammonium glycyrrhizinate protected cardiomyocytes in ISO-induced MI model by inhibiting LTCCs and reducing oxidative stress, with the similar mechanism as glycyrrhizic acid [76].Above results suggested derivatives of glycyrrhizic acid may be a promising drug for treating cardiovascular disease.

Calenduloside E
Aralia elata (Miq.)Seem is a traditional Chinese medicinal plant used for treating arrhythmia, diabetes and coronary heart disease.Calenduloside E (17, Fig. 5), a pentacyclic triterpenoid saponin from Aralia elata (Miq.)Seem., has the anti-MI, anti-hypoxia and anti-endothelial injury activity [77].Calenduloside E was verified the protection effect by suppressing calcium overload though restoring the expression of calcium transporters, such as SERCA, α 1 C(Cav1.2),RyR2 and NCX, and regulating the calcium transients in MI/RI rats [77].Further research showed that Calenduloside E enhanced the interaction between LTCCs and Bcl2-associated athanogene 3 (BAG3), specifically by inhibiting the α 1 C (Cav1.2) and α 2 δ subunits of LTCCs, restoring the interaction between BAG3 and α 1 C to alleviated MI/R injury [77].In addition, the mechanism of calenduloside E has been found to be similar to that of nisoldipine, a dihydropyridine calcium channel blocker, suggesting that calenduloside E has the potential to be developed as an LTCCs antagonist.

Alkaloids Sinomenine
Sinomenine (18, Fig. 6), a major bioactive alkaloid from Sinomenium acutum, has protective effects on cardiocerebrovascular system [78].Sinomenine protected against the oxygen-glucose deprivation-reperfusion induced neurotoxicity in PC12 cell, and improved functional recovery in cerebral ischaemia rats [79].Specifically, sinomenine inhibited L-type calcium currents, decreased [Ca 2+ ]i induced by acidification, and reduced ASIC1a currents, which directly induced Ca 2+ entry in rat cultured cortical neurons [79].The sinomenine is expected to be applied in the prevention and treatment of stroke.
In addition, the neuroprotective effects of ligustrazine have also been reported [83,84].Ligustrazine showed protective effects on hippocampal neuron cells mainly by inhibiting I Ca,L and reducing intracellular calcium concentration [83].Moreover, the protection of ligustrazine on SH-SY5Y human neuroblastoma cells by inhibiting LTCC were reported [84].

Berberine and coptisine
Berberine and coptisine (20-21, Fig. 6) are active alkaloids widely existing in Coptis species, which have anti-tumor, anti-microbial and cardio-cerebrovascular protection effects [85].Berberine treatment inhibited LTCCs by decreasing the expression of α 1 C subunit and the intracellular Ca 2+ level in smooth muscle cells of streptozotocin-induced diabetes rats [86].Berberine enhanced the neuroprotective effect of verapamil in sporadic dementia of Alzheimer's type rats induced by intracerebroventricular streptozocin by inhibiting of P-gp efflux and regulating calcium homeostasis [87].Berberine exerted the positive inotropic effect on left ventricular myocytes of rat heart by enhancing the Ca 2+ influx [88].In addition, bromibenzyltetrahydroberberine (CPU86035), a tetrahydroberberine derivative, strongly inhibited LTCCs in single ventricular myocyte of guinea pig, which can be used in the treatment of myocardial infarction [89].Therefore, the effects of berberine and its derivatives may provide therapeutic strategies for calcium channel diseases.
Coptisine relaxed abnormal contracted mouse airway smooth muscle (ASM) by eliminating LTCCs and and regulating intracellular Ca 2+ concentration, and exhibited the similar calcium antagonism as nifedipine [90].

Ibogaine
Ibogaine (22, Fig. 6), an indole alkaloid isolated from the root bark of the African shrub Tabernanthe iboga with a long history usage as medicinal agent to treat drug abuse in local aera [91].The study revealed that its anti-addictive effect of ibogaine was associated with the inhibition of Cav 1.2 channel in guinea pig cardiomyocytes [92].

Cinobufagin and bufalin
Chan Su, a Chinese medicine made from the dried white secretions of Chinese toads (Bufo melanostictus Schneider or Bufo bufo gargarizans Cantor) [93].Cinobufagin and bufalin (23-24, Fig. 7), two natural bufaldienolides Fig. 6 Chemical structures of alkaloids, including sinomenine, ligustrazine, berberine and coptisine from Chan Su, have been reported the cardioprotective effect by inhibiting LTCCs.Cinobufagin induced certain electrophysiological changes in the properties of I Ca,L in a concentration-dependent manner [93].Bufalin inhibited the I Ca,L , and contractility in rat ventricular myocytes induced by Bay K8644, an LTCCs agonist [94].

Phenylpropanoids Cinnamaldehyde
Cinnamaldehyde (25, Fig. 7), an active natural ingredient from the Cinnamomum tamala (Bauch.-Ham.), has been used in treating cardiovascular diseases [95].Cinnamaldehyde exhibited complex effects on rat aortic rings and isolated mouse hearts by activating the chemosensory cation channel TRPA1 as well as inhibiting LTCCs.The inhibitory action of cinnamaldehyde on I Ca,L in both ventricular cardiomyocytes (VCM) and vascular smooth muscle cells(VSMC) was obtained and these effects were similar to those of classic LTCCs blocker verapamil [95].

Others Salidroside
Salidroside (26, Fig. 8) is a phenylethanoid derivative in Rhodiola rosea L. and has obvious hypoglycemic effect in diabetes.Recently, studies have explored the antihypertensive mechanism of salidroside in diabetic vascular complications.Salidroside dilated the cerebral arteries of diabetic rats, but could not recover to the normal level, and had no diastolic effect on the cerebral arteries of normal rats.In this process, calcium current density, the protein and mRNA expressions of α 1 C subunit at diabetic rats were inhibited by salidroside [96].Salidroside also showed the protection hippocampal neurons against hypoxic-induced injury based on inhibiting LTCCs and reducing the mRNA expression levels of Cav1.3 and NMDAR1 to alleviate the intracellular calcium overload [97].

Crocin
Crocin (27, Fig. 8), a major bioactive substance from Crocus sativus L., have various effects on cardio-cerebrovascular system, including anti-oxidation, anti-MI and MI/ RI [98].In terms of anti-MI, crocin exerted negative inotropic effects on myocardium, reduced oxygen consumption and protected myocardium cells by inhibiting I Ca,L and reducing Ca 2+ influx [99].

LTCCs activators from natural products
There are few studies on LTCCs activators in TCM.Studies have shown that quercetin (28, Fig. 9, 10 μM) activated Cav1.2 channel current (I Ca1.2 ), negative shifted the steady-state inactivation curve and slowed recovery rate from inactivation in rat tail artery [100].However, the electrophysiological features of quercetin on I Ca(L) were different from Bay K 8644, a known Ca 2+ channel agonist.The in-depth research showed that the ineffective concentrations of quercetin (0.1 and 0.3 μM) inhibited the max response induced by Bay K 8644, indicating that low dosage of quercetin may restricted the LTCC reaction stimulated by Bay K 8644 [101].Another study showed that quercetin induced insulin secretion by directly activating LTCCs in insulin-secreting cell line INS-1, which has potential for controlling type 2 diabetes [102].Myricetin (29, Fig. 9), a analogue of quercetin, exerted the similar activatation on L-type Ca 2+ channel with (S)-(-) Bay K 8644, by slowing down the activation kinetics and stabilizeing the channel in its inactivated state [103,104].
The regulatory ability of flavonoid constituents on Cav1.2 channels were summarized [105].Twenty-four flavonoids were conducted to measure their effects on I Ca1.2 in rat tail artery myocytes with patch-clamp Method.As a result, the effect of flavonoids on calcium channels is voltage dependent, six compounds including quercetin (28), myricetin (29), isorhamnetin (30), luteolin (31), apigenin (32) and kaempferol (33) enhanced the I Ca1.2 with the EC 50 of ranging between 2.9 and 16.0 mM (Fig. 9).The affinity of quercetin to Cav1.2 was 3 times that of myricetin, and the effect of quercetin was significantly better than that of myricetin.The hydroxyl position and the double bond between C2 and C3 in skeleton of flavonoids were the important determinants for predicting the activity of flavonoids on calcium channels by molecular modelling method [105].

Discussion
LTCCs is an important voltage-gated channel that are responsible for regulating intracellular calcium balance and participating in a variety of human diseases, which has been considered as the potential therapeutic target.Abnormal LTCCs expression is closely related to the progression of cardiovascular, neurological and psychological diseases.Correcting calcium homeostasis disorders may be successful therapeutic strategies in the treatment Fig. 9 Chemical structures of quercetin, myricetin, isorhamnetin, luteolin, apigenin and kaempferol above diseases or delay the progression of diseases [106].Many non-natural LTCCs antagonists have been used in clinical practice for decades, such as nimodipine, diltiazem and verapamil [11].Multiple types of natural LTCCs antagonists from TCM, including polyphenols, flavonoids, terpenoids, alkaloids, steroids and phenylpropanoids were summarized in this review (Fig. 10).Most natural LTCCs antagonists mentioned in the article were isolated from herbs (92.6%), but part of them, such as Cinobufagin and bufalin, were obtained from animal (7.4%) [93,94].Animal medicine was one of the main sources of TCM, that has received enough attention in TCM research.However, it is necessary to pay attention to the sustainable utilization of resources and the protection of ecological environment during the exploration of new animal medicine resources [107].Actually, these natural LTCCs antagonists have been still in laboratory stage and not used in clinic so far.
Research revealed that non-selective calcium channel antagonists were dose-limited clinically by vascular effects and were prone to cause adverse reactions such as peripheral oedema, headache and dizziness [108].Currently, the selective Cav1.3 blockers had significant therapeutic effects but without those vascular side effects of non-selective LTCCs blockers [43].Therefore, specific LTCCs antagonists need to be developed in future.Furthermore, the LTCCs were new promising targets for many diseases, such as drug-addiction [33,34], depressive disorder [109], age-related macular degeneration-retinal pigment epithelium (AMD-RPE) [110], intrauterine growth restriction [111], local infiltration analgesia [112], myalgia [113], which expanded the scope of application.In addition, calcium channel blocker (CCB) have a short plasma half-life especially in rodents and show high first-pass metabolism upon oral application [114].The dosage of CCB should be tightly noticed as the CCBs lose specificity for their specific receptors and can show all the manifestations of toxicity such as bradycardia, peripheral vasodilation, and hypotension in high concentrations [115].

Conclusions
LTCCs is a promising target to develop as its involvement in various heritable complex diseases..Nowadays, many natural products from TCM have been reported the inhibitary activity on LTCCs, but their molecular characteristics, intricate mechanisms, and shortage of clinical evidence limits their development and usage.

Fig. 1
Fig. 1 Subunit structure of LTCCs.It consists of α 1 , α 2 , δ, β and γ subunits, and the α 1 subunit consists of four domains with six fragments in each domain (S1-S6).The positively charged S4 responds to the membrane potential change by transferring to the pore domain via the cytoplasmic S4-S5 connector.The motion of S4 is guided by the negatively charged interaction provided by the S1-S3

Fig. 2
Fig.2The LTCCs antagonists from natural products.The excitation-contraction coupling process begins with the entry of Ca 2+ into the cell through LTCCs, followed by the triggering of Ca 2+ release on SR by RyR2, and ultimately triggering intracellular Ca 2+ concentration mediated contraction.Subsequently, Ca 2+ in the cytoplasm is brought back to SR by SERCA and transported back to extracellular space via NCX (black arrow).Activated CaMKII induces stimulatory actions by phosphorylating major Ca 2+ homeostatic proteins, activating I Ca,L (mediated by the Thr498 terminal of α and β 2a subunits), phospholamban to increase cytosolic Ca 2+ uptake by the SR, and RyR to increase SR Ca 2+ release (blue arrow).Some active ingredients in TCM can antagonize LTCCs through various mechanisms.Inhibition of LTCCs subunits (α 1 C, α 1 D, α 2 δ), CaMKII phosphorylation and I Ca,L reduced the release of Ca 2+ from the sarcoplasmic reticulum

Table 1
Classification and pharmacology of LTCCs

Table 2
Pharmacological effect and mechanism of natural products of TCM on LTCCs